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Pan-Genome Analysis of the Genus Finegoldia Identifies Two Distinct Clades, Strain-Specific Heterogeneity, and Putative Virulence Factors

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Pan-Genome Analysis of the Genus Finegoldia Identifies Two Distinct Clades, Strain-Specific Heterogeneity, and Putative Virulence Factors Supplementary Information Pan-genome analysis of the genus Finegoldia identifies two distinct clades, strain-specific heterogeneity, and putative virulence factors Holger Brüggemann, Anders Jensen, Seven Nazipi, Hüsnü Aslan, Rikke Louise Meyer, Anja Poehlein, Elzbieta Brzuszkiewicz, Munir Al-Zeer, Volker Brinkmann, Bo Söderquist Content: Figure S1: Phylogenetic tree of 16S rDNA sequences extracted from draft genomes of Finegoldia Figure S2: Phylogenetic tree of 16S rDNA sequences of Finegoldia isolates extracted from GenBank Figure S3: Predicted genomic islands in Finegoldia genomes Figure S4: Phylogenetic tree of all putative CAMP factors of Fingoldia strains Figure S5: CAMP reaction of Finegoldia strains at elongated cultivation time Figure S6: Insertional inactivation of camp1 in strain CCUG54800 Table S1: Bidirectional blast of all coding sequences of 12 Finegoldia genomes Table S2: CRISPR/cas systems in Finegoldia sp. Tables S3A-S3L: Predicted genomic islands in the genomes of F. magna and “F. nericia” Table S4A-S4E: Blast results showing homologs of host-interacting factors in 12 Finegoldia genomes Table S5: Finegoldia sp. strains tested in the RapIDTM ANA II system Figure S1: Phylogenetic tree of 16S rDNA sequences extracted from draft genomes of Finegoldia The analysis involved 16 nucleotide sequences. For one genome, F. magna ALB8, the 16S rDNA sequence is incomplete. There is an overall high similarity (in average 99%) between the 16S rDNA sequence of strains of F. magna and “F. nericia”: there are 5 to 9 SNPs in the 16S rDNA of “F. nericia” strains compared to the cluster of F. magna strains, including the reference strain F. magna ATCC29328. The phylogenetic analysis was done with MEGA7. The evolutionary history was inferred using the Neighbor-Joining method. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (250 replicates) are shown next to the branches. The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Maximum Composite Likelihood method and are in the units of the number of base substitutions per site. F magna GED7760A F magna 08T492 F magna ACS-171-V-Col3 F magna SY403409CC001050417 F magna ATCC29328 56 KR232883.1 Finegoldia sp. S393 KC999388.1 Finegoldia sp. S8 F7 KM461968.1 Finegoldia sp. feline oral taxon 341 strain 7209 F. magna JN809779.1 Finegoldia sp. BV3C29 37 JX103994.1 Finegoldia sp. MVA12 JX104040.1 Finegoldia sp. S4-BM10 F magna 09T408 6 KC010025.1 Uncultured Finegoldia sp. clone 5 30 33 F magna BVS033A4 JX103989.1 Finegoldia sp. MVA2s 15 68 F magna 07T609 17 F nericia T160124 KF007163.1 Finegoldia sp. S9 AA1-5 2 KP944179.1 Finegoldia magna strain 1309-13310 1 F nericia 12T273 AB691574.1 Finegoldia magna strain JCM 6491 5 47 NR 041935.1 Finegoldia magna strain CCUG 17636 48 NR 113383.1 Finegoldia magna strain JCM 1766 AY169413.1 Finegoldia magna clone 1-9 15 F nericia 12T306 8 24 AB691573.1 Finegoldia magna strain JCM 6489 “F. nericia” 30 F nericia CCUG54800 EU704222.1 Uncultured Finegoldia sp. clone KLOND11 17 EU530177.1 Uncultured Finegoldia sp. clone M1-26 EU704220.1 Uncultured Finegoldia sp. clone KLOND09 6 JN809765.1 Finegoldia sp. BV3Pr2 EU704207.1 Uncultured Finegoldia sp. clone KLONB11 60 JX103990.1 Finegoldia sp. MVA6 59 F nericia 09T494 45 GQ179678.1 Uncultured Finegoldia sp. clone VE49H01 19 F nericia 12T272 EU704205.1 Uncultured Finegoldia sp. clone KLONB08 F nericia ATCC53516 EU704209.1 Uncultured Finegoldia sp. clone KLONC02 EU704214.1 Uncultured Finegoldia sp. clone KLONC12 42 EU704218.1 Uncultured Finegoldia sp. clone KLOND05 EU704211.1 Uncultured Finegoldia sp. clone KLONC06 EU704198.1 Uncultured Finegoldia sp. clone KLONA10 64 EU704195.1 Uncultured Finegoldia sp. clone KLONA06 F nericia T151023 77 EU704206.1 Uncultured Finegoldia sp. clone KLONB10 76 EU704210.1 Uncultured Finegoldia sp. clone KLONC03 0.00050 Figure S2: Phylogenetic tree of 16S rDNA sequences of Finegoldia isolates extracted from GenBank All deposited 16S rDNA sequences assigned to Finegoldia were taken from GenBank that had a >85% overlap with the 16S rDNA sequence (rrnA) of ATCC29328. 47 sequences were compared. The phylogenetic analysis was done with MEGA7. The evolutionary history was inferred using the Neighbor-Joining method. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (250 replicates) are shown next to the branches. The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Maximum Composite Likelihood method and are in the units of the number of base substitutions per site. 07T609 08T492 09T408 09T494 12T272 12T273 12T306 T151023 T160124 CCUG54800 ATCC 53516 ATCC 29328 Figure S3: Predicted genomic islands in Finegoldia genomes All analyzed 12 Finegoldia genomes contain predicted genomic islands, but with strain-specific differences. The color depicts the prediction method: blue, IslandPath-DIMOB; orange, SIGI- HMM and dark red, integrated. All genes found in these islands are shown in table S3. CAMP1 CAMP2 CAMP3 CAMP4 Figure S4: Phylogenetic tree of all putative CAMP factors of Finegoldia strains Each strain possesses a least two distinct CAMP factors whose genes are in direct vicinity. Four out of seven “F. nericia” strains possess two additional CAMP factors; again, the genes are located next to each other. The tree was created in MEGA7, using the Maximum Likelihood method. T 0 1 0 0 1 0 9 2 7 8 5 9 T T T T 1 T 4 2 6 4 0 4 9 7 0 9 2 0 4 2 9 2 3 8 S. aureus S. aureus T C 1 C 1 1 6 5 2 2 0 4 T T 1 8 2 3 2 0 7 0 4 0 3 6 Figure S5: CAMP reaction of Finegoldia sp. strains at elongated cultivation time TSA agar plates with 5% sheep blood was used. Staphylococcus aureus is inoculated in the middle streak. A positive CAMP test is indicated by complete erythrocyte lysis at the interface of the Finegoldia sp. and the S. aureus streaks. All three F. magna (in red) strains and two out of seven “F. nericia” (in blue) strains showed a strong positive CAMP reaction after five days of anaerobic incubation. One strain (CCUG54800) is negative in the CAMP reaction (see also Figure S6). “F. nericia” CCUG54800 insE camp1 insO camp2 camp1 camp2 “F. nericia” 12T273 Figure S6: Insertional inactivation of camp1 in strain CCUG54800 The genomic regions containing the camp1 and the camp2 gene in the “F. nericia” strains CCUG54800 and 12T273 are shown. A transposase-encoding mobile element is inserted in the 5’ end of camp1 in strain CCUG54800. Table S1: Bidirectional blast of all coding sequences of 12 Finegoldia genomes The program ProteinOrtho (version 4.26) was used with the following parameters: blast=blastp v2.2.24, E-value=1e-10, alg.-conn.=0.1, coverage=0.5, percent_identity=25, adaptive_similarity=0.95, inc_pairs=1, inc_singles=1, selfblast=1, unambiguous=0) The Prokka annotation is shown. Highlighted in grey are the CDS with orthologs in all 12 genomes. #species proteins alg.-conn. Annotation ATCC29328 ATCC53516 07T609 08T492 09T408 09T494 12T272 12T273 12T306 CCUG54800 T151023 T160124 #12 21125 - 1842 CDS 1817 CDS 1698 CDS 1796 CDS 1677 CDS 1827 CDS 1773 CDS 1840 CDS 1712 CDS 1906 CDS 1570 CDS 1667 CDS 12 12 1 Chromosomal replication initiator protein Dna ATCC29328_00010 ATCC53516_11280 07T609_16840 08T492_17540 09T408_16640 09T494_01290 12T272_17700 12T273_00730 12T306_00150 CCUG54800_18780 T151023_00340 T160124_16570 12 12 1 DNA polymerase III subunit beta ATCC29328_00020 ATCC53516_11270 07T609_16850 08T492_17550 09T408_16650 09T494_01300 12T272_17710 12T273_00740 12T306_00160 CCUG54800_18790 T151023_00350 T160124_16580 12 12 1 ribosome-associated protein ATCC29328_00030 ATCC53516_11260 07T609_16860 08T492_17560 09T408_16660 09T494_01310 12T272_17720 12T273_00750 12T306_00170 CCUG54800_18800 T151023_00360 T160124_16590 12 12 1 DNA replication and repair protein RecF ATCC29328_00040 ATCC53516_11250 07T609_16870 08T492_17570 09T408_16670 09T494_01320 12T272_17730 12T273_00760 12T306_00180 CCUG54800_18810 T151023_00370 T160124_16600 12 12 1 hypothetical protein ATCC29328_00050 ATCC53516_11240 07T609_16880 08T492_17580 09T408_16680 09T494_01330 12T272_17740 12T273_00770 12T306_00190 CCUG54800_18820 T151023_00380 T160124_16610 12 12 1 DNA gyrase subunit B ATCC29328_00060 ATCC53516_11230 07T609_16890 08T492_17590 09T408_16690 09T494_01340 12T272_17750 12T273_00780 12T306_00200 CCUG54800_18830 T151023_00390 T160124_16620 12 12 1 DNA gyrase subunit A ATCC29328_00070 ATCC53516_11220 07T609_16900 08T492_17600 09T408_16700 09T494_01350 12T272_17760 12T273_00790 12T306_00210 CCUG54800_18840 T151023_00400 T160124_16630 12 12 1 Putative anti-sigma factor antagonist ATCC29328_00080 ATCC53516_11210 07T609_16910 08T492_17610 09T408_16710 09T494_01360 12T272_17770 12T273_00800 12T306_00220 CCUG54800_18850 T151023_00410 T160124_16640 12 12 1 serine-protein kinase RsbW ATCC29328_00090 ATCC53516_11200 07T609_16920 08T492_17620 09T408_16720 09T494_01370 12T272_17780 12T273_00810 12T306_00230 CCUG54800_18860 T151023_00420 T160124_16650 12 24 0.167 RNA polymerase sigma factor SigF ATCC29328_00100,ATCC29328ATCC53516_16000,ATCC5351607T609_08200,07T609_108T492_08050,08T492_17 09T408_08280,09T408_109T494_09240,09T494_012T272_07000,12T272_12T273_11890,12T273_12T306_10120,12T306_CCUG54800_18870,CCUG548 T151023_08580,T151023_ T160124_08490,T160124_16660 12 12
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